Theophylline is a bronchodilator and respiratory stimulant that is effective in the treatment of acute and chronic asthma, Cheyne-Stokes respirations, and apnea/bradycardia episodes in newborn infants. It is also used as an adjunct in the treatment of congestive heart failure and acute pulmonary edema. The most important current use of theophylline is as a prophylactic agent for controlling the symptoms of chronic asthma.
The therapeutic range for theophylline is relatively narrow, around 10-20 .mu.g/ml (55-110 .mu.M). Since the metabolism of theophylline varies from individual to individual, close monitoring is needed in order to avoid harmful side-effects caused by overdoses. The need for a quick, easy-to-use device which can monitor the narrow theophylline therapeutic range has been recognized for some time. Such devices have to cover not only the therapeutic range, but also higher and lower levels; in cases of toxicity, levels as high as 60 .mu.g/ml (330 .mu.M) may be encountered.
There are many current methods for measuring theophylline, including chromatographic and immunoassay procedures. In any assay method, advantage is taken of the knowledge that theophylline can be determined by measuring its inhibition of alkaline phosphatase. However, when assaying human biological fluids in this manner, endogenous alkaline phosphatase can effect the assay and render results inaccurate.
In EP 188372, a method of eliminating the effect of endogenous alkaline phosphatase is described. Theophylline is a very potent inhibitor of certain isozymes of alkaline phosphatase, particularly that obtained from bovine liver. At a pH less than 9, this isozyme exhibits greater catalytic activity than isozymes present in human serum. When bovine liver alkaline phosphatase and a substrate, p-nitrophenylphosphate, are present in a layered strip along with a buffer to give the lower pH, the application of a sample to the strip results in an enzyme activity at pH 9 or less, and gives a color intensity that is inversely proportional to the concentration of theophylline in the sample. The color is read from the back of the strip using a reflectance photometer.
A modified system is described in EP 258034, in which the buffer is in a porous spreading zone. EP 258035 describes another modification, in which a greater quantity of isoenzyme is employed. Both of these modifications are intended to minimize the effect of endogenous alkaline phosphatase, among other objects.